Salmonella enterica has been increasingly linked to outbreaks involving consumption of fresh produce. Although researchers have identified genes whose products are involved in mediating S. enterica–plant interactions, the use of various experimental approaches, serovars, and plant types has generated variable and conflicting data. The purpose of this study was to determine whether conditions under which inocula are prepared for in vitro plant interaction studies influence the outcome of these studies. Seven S. enterica serovars were grown in media that differed in salinity and physical state with incubation at 25 or 37°C. These cultures were then used to inoculate red leaf lettuce, and adherent microbes were subsequently recovered. Although all Salmonella serovars were influenced by inoculum preparation conditions, the amount of variation differed. Analysis of pooled serovar data revealed that inocula prepared from either agar plates or biphasic cultures had higher levels of interaction with red leaf lettuce than those prepared from broth cultures. Incubation at 37°C enhanced adherence after 30 s or 5 days of contact time, and adherence after 1 h of contact time was increased in low-salt medium. Broth inoculum cultures were highly influenced by medium salinity and incubation temperature, whereas plate and biphasic inoculum cultures were only minimally affected. Therefore, inocula prepared from bacteria grown on plates or in biphasic culture would be most suitable for evaluation of strategies used to interfere with plant-Salmonella interactions. However, pooled data mask serovar-specific responses, and care should be taken when extrapolating these findings to individual serovars. The previous association of a serovar with outbreaks involving leafy greens was not correlated with levels of interaction with red leaf lettuce, suggesting that the occurrence of these serovars in or on these commodities does not reflect their fitness in the plant environment.

  • Plate-grown Salmonella inocula had increased interactions with red leaf lettuce.

  • Salmonella serovars were not equally influenced by inoculum growth conditions.

  • Inoculum growth temperature influenced early (30 s) and late (5 days) adherence to lettuce.

  • Medium salinity influenced Salmonella adherence after 1 h of contact time.

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